The trichomonad parasite, Trichomonas vaginalis, causes a major sexually transmitted disease, trichomoniasis, in humans, an infection of substantial medical importance. T vaginalis is the most common cause of vaginitis in women and has been linked to several major pathological symptoms such as severe vaginal inflammation and irritation, infertility, preterm delivery, low-birth-weight infants, invasive cervical carcinoma and increased intrauterine transmission of cytornegalovirus and also susceptibility to human immunodeficiency virus (HIV) infection. The ability of parasites to adhere to host cells plays an integral role in establishing infections. Such interactions may be mediated by cell surface glycoconjugates. The long term objective of this project is to use highly purified parasite glycoconjugate products to determine the role of cell-bound and free lipophosphoglycan (TV-LPG) upon T vaginalis infection pathology, andimmunity. The parasite possesses novel LPG-like glycoconjugates (3 X 106 copies/parasite) anchored on the cell surface via an inositol phosphoceramide, distinct from any other GPI anchor reported so far. The experimental data suggest that TV-LPG is involved in adhesion of T vaginalis to HeLa cells through a specific receptor-ligand interaction. The water soluble TV-LPG related glycan (TV-GL) purified from conditioned media of T vaginalis may also be involved in T vaginalis pathogenesis. The specific aims of the project are: 1) to establish in vitro cultures of human vaginal epithelial cells (HVECs) to study the involvement of the LPG-like glycoconjugates in adhesion of T vaginalis to HVECs and also examine the hormonal effects of adhesion of T vaginalis; II) to structurally characterize TV LPG and clarify its relationship to the water soluble TV-GL; III) to produce monoclonal antibodies (mAbs) against LPG and its fragments, to examine which LPG epitopes are expressed on the surface of the live parasite; and IV) to determine if mAbs specific for surface expressed LPG epitopes inhibit adhesion to HVECs and are cytotoxic to the parasite in vitro. The glycoconjugates from cultured parasites are analyzed by chemical and enzymatic digestion in combination with HPLC, Glyko-FACE, GC/M[S, MS, MALDI-TOF MS andNMR. HVECS are being cultured as we have established for bovine "vaginal' epithelial cells to study bovine trichomonad pathogenesis, and will be exposed to hormone to study the binding of T vaginalis to HVECs. These biological and chemical studies will provide essential knowledge of the structure/function relationships of TV-LPG and will lead to a better understanding of the molecular mechanisms of pathogenesis involved in host-parasite interactions. In addition, these findings will contribute to the development of effective diagnosis and therapies of human trichomoniasis.